Bioassay-guided separation led to identification of bostrycin1, which was a potentially promising anti-tumor drug candidate, together with other four anthraquinone 2-5 derivatives from liquid culture of endophytic Nigrospora sp. BM-2 isolated from Saccharum arundinaceum Retz. In order to obtain high bostrycin production, three single-factor experiments and a L9(34) orthogonal matrix experimental design have been employed in the cultivation of Nigrospora sp. BM-2. The optimal fermentation medium were glucose 20.0 g/L, yeast extract 8.0 g/L, MgSO4 0.05 g/L, KH2PO4 0.01g/L, NaCl 0.5 g/L. After 9 days of incubation under shake flask conditions at 120 rpm and 28 °C in optimal media, the highest amount of bostrycin was 86.25 ± 7.23 mg/L in culture broth. Furthermore, enzymatic inhibitor as regulate factor in the biosynthesis of bostrycin was attempted to give adverse effects.